SHN3多克隆抗体纯化与鉴定

Purification and Identification of Polyclonal Antibodies against SHN3

目的:利用SHN3免疫特异肽段亲和层析纯化抗SHN3多克隆抗体。方法:将制备多克隆抗体SHN3免疫原片段(99bp)划分为4段互相重叠等长的基因片段(48bp),构建原核表达重组质粒并转化E.coil的BL21(DE3)菌株。IPGT诱导重组蛋白表达,Western blotting筛选并人工合成与抗体免疫结合特异性片段,亲和层析纯化抗体,Western blotting检测纯化结果。结果:成功地筛选到SHN3免疫原特异肽段part-3,以亲和层析的方式纯化多克隆抗体,纯化后免疫特异性提高。结论:亲和层析的方式纯化得到的SHN3多克隆抗体抗体,有效地提高免疫特异性,为后续SHN3蛋白功能研究和应用奠定了基础。

Objective：To purify polyclonal antibodies against SHN3 by affinity chromatography with immunologic specificity peptide frag- ment. Method：SHN3 immunologic specificity gene （90bp）was divided 4 equal length parts（48bp）. And subcloned into prokaryotic ex- pression vector Pmal - C2. These recombinant vectors were transformed into E. coliBL21 （ DE3 ）. Recombinant peptide expression were in- duced by IPTG. Western blotting method was used to screen the best immunologic specificity peptide fragment. The polyclonal antibodies was purified by affinity chromatography and checked by Western blotting. Result：The immunologic specificity peptide fragment part -3 of SHN3 was screened successfully, and the SHN3 polyclonal antibodies were purified with this peptide by affinity chromatography. The affini- ty chromatography of SHN3 polyclonal antibodies was improved significantly. Conclusion： Affinity chromatograph for SHN3 polyclonal anti- bodies purification was successfully established and immunologic specificity was significantly improved. The method laid a foundation for subsequent SHN3 research and application.